The present invention relates generally to photometric devices and apparatuses for spectroscopic analysis of small absolute amounts of analyte in solution, and more particularly to a flow cell assembly using an amorphous fluoropolymer material for light conduction through the flow cell.
Light absorption detectors for high performance liquid chromatography (xe2x80x9cHPLCxe2x80x9d), capillary liquid chromatography (capillary LC or xe2x80x9cCLCxe2x80x9d), capillary electrophoresis (CE) and capillary electrochromatography (CECxe2x80x9d) generally include four basic components; a light source, a means for selecting range of wavelengths, a flow cell and at least one light sensor that measures the amount of light being transmitted through the flow cell. The apparatus may be disposed with parallel optical paths as a double beam instrument. The flow cell is typically in the form of a hollow tube through which a sample to be analyzed and the light are passed. Also these four basic components are typically configured to be in a predetermined arrangement with respect to each other. With these kinds of detectors, it is known that when a sample to be analyzed passes through the flow cell, the amount of light transmitted through the flow cell decreases in accordance with Beer""s law.
In conventional flow cells, light is typically passed through the flow cell in one of two fashions, along the long axis of the hollow tube or perpendicular to the long axis. In either case, i.e. when the light is parallel to the long axis or introduced perpendicular thereto, the detector or light sensor output is usually expressed in terms of absorbance, which is proportional to the sample concentration and the pathlength. Thus, the longer the pathlength, the larger the detector output signal should be for a given sample concentration. For conventional flow cells, however, the light striking the lateral wall of the flow cell is partially lost due to absorption and scattering at the wall. This lost light reduces the light energy throughput of the flowcell thereby causing an increase in the noise in the output signal of the detector.
The lateral dimension or diameter of the flow cell could be increased to reduce the fraction of light striking the lateral wall, but this increases the volume of the flow cell. Frequently, however, the quantity of the sample is limited, thus the optimum cell has a minimum volume implying a small cross-section or diameter. In addition, a larger cell volume also has the effect of spreading out or dispersing the sample peak and causing a loss in chromatographic resolution. Thus, as a practical matter the foregoing effects limit the pathlength and lateral dimension of conventional flow cells.
There is described in U.S. Pat. No. 5,608,517 a flow cell having a flow passage that is coated with a polymer having an index of refraction lower than that of common chromatography solvents, e.g. water. With such a flow cell, the light being directed into the flow cell is internally reflected or piped down the length of the flow passage. It is difficult, however, with such a flow cell to obtain consistent well adhered layers of the polymer on the inner wall of the housing for the flow cell. Thus, it is possible for the polymer layer to become detached from the housing inner wall or delaminated therefrom. This delaminating effect leads to distortion of the fluid flow channel causing distortion of the optical path and the fluid flow in the flow cell, as well as creating a condition whereby the fluid could flow between the polymer and the housing.
There also is described in U.S. Pat. No. 5,184,192, owned by the assignee of the present invention, a flow cell having an inner wall formed of an amorphous fluoropolymer having a refractive index less than the refractive index of common chromatography solvents, e.g. water. Although the flow cells described therein are capable of yielding a cell with a long pathlength, the process required for making such a flow cell, and the resultant flow cell, may not be suitable for particular applications.
Moreover, the apparatuses or systems using either of these referenced flow cells must be arranged so the flow cell is precisely located with respect to the other components constituting the apparatus or system. That is, these flow cells must be precisely located in the optical path between the light source and the light sensor or detector. The criticality of the location of known flow cell(s) is the result of the need for continual, optimal alignment, which yields greater reliability and greater analytical reproducibility. Additionally, it is generally preferred to minimize the length of the flow channel between for example the chromatography column and the flow cell, which is necessarily limited by construction and design of such apparatuses or devices. Thus, even though the above described flow cells may be capable of achieving a long path length, they are typically located with respect to other components of the detection device or apparatus in the same fashion as conventional flow cells, which do not generally optimize placement from a chromatography standpoint.
A flow cell described in U.S. Pat. No. 4,867,559 includes a cladding liquid passed through a capillary in order to coat the interior bore with a low refractive index fluorocarbon. The fluorocarbon is disclosed as being a viscous, inert, immiscible, nonwetting material, such as fluorinated oil generally available under the tradenames Fluorinert(copyright) or Krytox(copyright). This arrangement, however, is extremely unsatisfactory in practice or use. This arrangement requires a complex fluid cladding handling mechanism, including pumps and plumbing, for delivering the cladding fluid to the capillary bore, and retrieving the excess fluid after the bore has been coated. Also, interfaces and seals must be implemented, which allow delivery of the cladding fluid to the bore while not interfering with delivery of the sample that is to be delivered to the bore subsequent to the coating by the cladding fluid. Further, the delivery and withdrawal of excess cladding fluid requires careful flow calibration, as the viscosity of the fluid and the small bore of the capillary can lead to irregular coating in the capillary. This adds considerably to the complexity and expense of the flow cell.
In addition, the cladding fluid is not bonded to the interior surface of the bore, thus it must be delivered shortly before the introduction of analyte in order to prevent settling of the cladding fluid inside the bore. Further, the cladding fluid may not fully coat the interior surface of the bore, leaving gaps in the cladding that leads to scattering of the optical signal and signal attenuation. Moreover, another problem that can result is the contamination of the sample with the cladding fluid, precluding the possibility of collection of pure fractions of the sample, which could possibly interfere with a downstream analytical technique, such as mass spectrometry.
The present invention provides a modular flow cell having a high optical throughput, a long optical path length and a small cross-section. The modular flow cell configuration includes remote ports or connections for liquid and light input, and liquid and light output. The modular flow cell can therefore be configured in a location optimized for chromatographic performance, in a chromatography apparatus or system. Novel methods are described for making the flow cell components, and for joining two materials or items together in fabrication of, for example, modular flowcell components. Such flow cells enable light to be guided axially along a sample filled tube or capillary, independent of the wall material of a transport tube. The modular flow cell of the present invention is optimally positioned relative to system components of a measuring apparatus or system and is easily installed as compared to prior art devices.
According to one aspect of the present invention the flow cell includes a flow cell body having two ends, each with a respective end interface secured thereto. The flow cell is configured to form a part of a modular flow cell assembly.
The flow cell body includes a channel having a through-aperture with an inner surface. A light guiding material, e.g. a transparent fluoropolymer material having a refractive index less than the refractive index of common chromatography solvents, is disposed proximate to the channel to form a light guiding through-aperture in the flow cell body. In a more specific embodiment, the flow cell body is included in a housing. The light guiding material in the flow cell body is, illustratively, one of a coating of a Teflon AF that is deposited onto the inner or exterior surface of the channel or is an extruded tubular member of Teflon AF that is in mechanical engagement with the channel. In any case, a hydraulic seal is established between the light guiding material and the boundary wall of the channel so the liquid sample does not bypass the through-aperture and flow along the exterior of the light guiding material on the inner surface. The channel is formed of any one of a number of materials that can provide the necessary mechanical strength and a fluid seal, and more particularly, includes materials which can develop a fluid seal at the interface between the flow cell body end and each end interface. Such materials include for example, polyetheretherketone (PEEK).
Each end interface includes an interface housing having a light transmitting passage and fluid flow passage disposed therethrough. The light transmitting passage is arranged in each end interface housing and the end interfaces are secured to the flow cell body so that one end of the light transmitting passage in each end interface is in optical communication with the through-aperture in the flow cell body. Each end of the flow cell body also is configured with an outwardly extending fluid channel that couples one end of the fluid flow passage in each end interface with the through-aperture of the flow cell body.
Each end interface includes a optical fiber as the light transmitting passage and a capillary tube, such as a quartz capillary tube, as the fluid flow passage. The interface housing in the illustrative embodiment is made of PEEK. In a more specific aspect of the present invention the interface housing is a PEEK over-mold in which is disposed the optical fiber and the capillary tubing to effectively form an integral structure. More particularly, the molding process is performed in such a manner that the PEEK intimately engages (e.g., adhesively bonds or frictionally engages) to the outer surface of the capillary tubing and the optical fiber, thereby establishing a strong mechanical connection and a hydraulic seal between the end member housing and each of the optical fiber and the capillary tubing. Additionally, the capillary tubing and/or optical fiber can include surface artifacts interior to the over-mold to improve the mechanical bonding with the housing. The free ends of the optical fiber and the capillary tubing are configured to extend outwardly from the end interface.
The flow cell according to the invention is integrated into an analyte measurement system for analyzing a fluid sample. The system includes a sensing device, a radiation source, a flow cell assembly with the flow cell according to the present invention, and interconnections optically interconnecting the flow cell assembly to each of the radiation source and the sensing device and fluidically interconnecting the flow cell with the fluid system(s). In such a system the flow cell assembly is capable of being located physically independent of, and thus isolatable (e.g., electrically isolated), from each of the radiation source and the sensing device. Thus, and in contrast to prior art devices, the flow cell assembly of the present invention does not have to be precisely located immediately adjacent to other system optical components to maintain a given optical configuration in order to reliably and reproducibly provide a signal output. In addition, because the flow cell assembly can be physically and electrically independent it can be configured modularly for easy replacement in the field.
The modularized flow cell assembly includes the flow cell as hereinabove described, mounted in a frame including optical fiber interconnections and capillary tubing interconnections. The fiber optic interconnections couple the free ends of the optical fiber extending outwardly from each end interface with optical ports provided in each of the radiation source and the sensing device. The capillary tubing interconnections are effected by any of a number of techniques known to those skilled in the art for fluidically interconnecting the free ends of the capillary tubing extending outwardly from the end interfaces to a fluid source and fluid sink, respectively.
The radiation source in the assembly incorporating the flow cell according to the invention, is a source of electromagnetic radiation that provides output in a predetermined fashion across a wide spectral range or a narrow band. In an illustrative embodiment, the radiation source is a light source including a deuterium lamp having spectral emissions encompassing the range from about 190nm to about 800nm.
The sensing device in the assembly according to the invention comprises a diffraction grating and photodiode array for sensing the light transmission through the fluid sample being analyzed, and for providing an output representative of that transmission.
In use, the fluid sample from the sample source (e.g., chromatographic column) flows through the flow passage or capillary tubing in one end interface and into the through-aperture in the flow cell body formed by the inner surface of the light guiding material. Fluid sample flows from the through-aperture, out through the flow passage or capillary tubing of the second end interface to a waste collection means or other downstream process. In this way, a static or flowing fluid sample is located in the through-aperture for analysis purposes. One end of the light transmitting passage or optical fiber extending out of one end interface is optically interconnected to the radiation source so the radiation or light therefrom passes through the light transmitting passage to the through-aperture in the flow cell body member. As described hereinabove this light is channeled by the light guiding material in the body so that the light traverses the cell substantially parallel to the long axis of the through-aperture. Correspondingly, the other end of the light transmitting passage or optical fiber extending out of the second end interface is optically interconnected to the sensing device so that the radiation or light that has traversed the cell exits the flow cell body through-aperture and passes into the sensing device.
Features of the invention include provision of a modular flow cell that is easily installed as a module in the context of system components, such as a radiation source and a sensing device. The flow cell and apparatus or system in which the flow cell is located is capable of being remoted a short distance from either the light source and light sensor or detector, as well as being electrically isolated therefrom. Such flow cells and measuring systems according to the invention are less costly and less difficult to manufacture in comparison to prior art devices. In addition, manufacturing and methods of use of the described flow cells and related apparatuses and systems are significantly simplified. The end interfaces are formed by a low cost, easily implemented over-molding process. The resultant flow cell is of a rugged, durable construction. The manner of construction allows one to easily manufacture flow cells having different pathlengths.